Biological lipid molecules consist of a polar structure (head group) and one or more non polar hydrocarbon chains. The hydrocarbon chains may contain one or more unsaturated C—C bonds. The number and position of these unsaturated bonds has a significant implication for biological activity.
It is known to use an Electrospray ion source in positive ion mode to ionise lipids and other biomolecules to form [M+H]+ singly charged molecular cations. The molecular cations may then be subjected to Collision Induced Dissociation (“CID”) so as to form a plurality of fragment ions. However, the resulting MS-MS mass spectra tend to be dominated by fragment ions associated with the polar head group of the lipid molecules and disadvantageously no structural information as to the position of the unsaturated bonds on the hydrocarbon chains is available.
The standard conventional approach involving using Collision Induced Dissociation is therefore unable to provide structural information as to the position of unsaturated bonds in lipids and other biomolecules. WO 2013/173642 (Sanford-Burnham Medical Research Institute) describes a high-throughput CID lipidomics technique.
Various attempts have been made to try to provide information relating to the location of e.g. double bonds in biological molecules.
US 2008/0296486 (Blanksby) describes a method for the determination of the position of unsaturation in a compound involving isolating a precursor ion and causing it to react with ozone. The position of unsaturation is determined using the mass difference between the isolated ion and an ozone-induced fragment ion.
H. T. Pham, A. J. Trevitt, T. W. Mitchell and S. J. Blanksby “Rapid differentiation of isomeric lipids by photodissociation mass spectrometry of fatty acid derivatives” Rapid Communications in Mass Spectrometry, 27(7), p. 805-815 (2013) discloses the derivatization of a structurally diverse suite of fatty acids as 4-iodobenzyl esters. Electrospray ionisation of these derivatives in the presence of sodium acetate yields abundant [M+Na]+ ions that can be mass selected and subjected to laser irradiation at a wavelength of 266 nm.
A. Devakumar, D. K. O'Dell, J. Michael Walker and J. P. Reilly “Structural Analysis of Leukotriene C4 Isomers Using Collisional Activation and 157 nm Photodissociation” American Society for Mass Spectrometry, vol. 19, no. 1 (2008) discloses the use of laser photodissociation as part of the structural characterisation of the two LTC4 isomers. Other laser photodissociation structural analyses are described in J. P. O'Brien and J. S. Brodbelt “Structural Characterisation of Gangliosides and Glycolipids via Ultraviolet Photodissociation Mass Spectrometry” Anal. Chem., 85, 10399-10407 (2013) and J. P. O'Brien, B. D. Needham, J. C. Henderson, E. M. Nowicki, M. Stephen Trent and J. S. Brodbelt “193 nm Ultraviolet Photodissociation Mass Spectrometry for the Structural Elucication of Lipid A Compounds in Complex Mixtures” Anal. Chem., 86, 2138-2145 (2014).
M. C. Thomas, T. W. Mitchell and S. J. Blanksby “Ozonolysis of Phospholipid Double Bonds during Electrospray Ionization: A New Tool for Structure Determination” J. Am. Chem. Soc. 128(1), p. 58-59 (2006) discloses ozonolysis of double bonds during the negative ion electrospray ionization of unsaturated phospholipids under conditions that produce a corona discharge. Ionic products of the ozonolysis are detected and characterised by mass spectrometry.
F. Hsu and J. Turk “Structural determination of sphingomyelin by tandem mass spectrometry with Electrospray ionization”, J. Am. Soc. Mass Spectrom., 11, p. 437-449 (2000) discloses forming alkaline metal adduct ions of sphingomyelin by Electrospray ionization.
S. L. Cook, O. L. Collin and G. P. Jackson “Metastable atom-activated dissociation mass spectrometry: leucine/isoleucine differentiation and ring cleavage of proline residues.” J. Mass. Spectrom., 44, p. 1211-1223 (2009) discloses extensive backbone fragmentation resulting in a-, b-, c-, x-, y- and z-type ions of singly and doubly charged peptide ions through their interaction with a high kinetic energy beam of argon or helium metastable atoms.
M. Green, K. Richardson, J. Brown and P. Murray “Investigation into UV photon-induced fragmentation in a RF confined ion guide using a commercial vacuum UV ionisation lamp” Poster ASMS May 2013 discloses using a UV lamp to fragment ions.
WO 2013/171495 (Micromass) discloses a mass spectrometer comprising a photoionisation device for excitation of reagent molecules within an RF ion guide.
WO 2013/021124 (Giuliani) discloses using a glow discharge lamp to fragment ions trapped in an ion trap.
Several of the known methods suffer from the serious disadvantage that the lipid molecules must first be derivatised or reacted with a separate reagent before analysis.
Several of the known methods suffer from the problem that the efficiency of the process is very low.
It is therefore desired to provide an improved method of mass spectrometry and in particular to provide an improved method of mass spectrometry to determine the location of double bonds in lipids.